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  • 1
    Electronic Resource
    Electronic Resource
    CYP2A6 genetic polymorphisms and liver microsomal coumarin and nicotine oxidation activities in Japanese and Caucasians (2000)
    Springer
    Archives of toxicology 73 (2000), S. 532-539 
    Details
    ISSN: 1432-0738
    Keywords: Key wordsCYP2A6 polymorphism ; Coumarin ; Nicotine ; Japanese ; Caucasian
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Genotypes of CYP2A6, namely CYP2A6 * 1 (wild-type), CYP2A6 * 2, and CYP2A6 * 3, were examined in liver DNA of 39 Japanese and 43 Caucasians using two-step polymerase chain reaction (PCR) methods. We first amplified a DNA fragment (1725 bp) located between near middle of exon 1 and end of exon 4 of the CYP2A6 gene and further amplified using a forward primer 't' or 'mut' (middle of exon 3) and a reverse primer 'E3R' (middle of intron 3) for the detection of CYP2A6 * 2-genetic polymorphism. The 1725 bp fragment was also used for the amplification between exon 3 and near middle of intron 3 of the CYP2A6 gene and the fragment thus obtained digested with XcmI or DdeI to detect and confirm the CYP2A6 * 2- and CYP2A6 * 3-types, respectively. Only one DNA sample from a Japanese origin (J18) was not amplified by CYP2A6-specific primers; liver microsomes from this individual had very low activity of coumarin 7-hydroxylation and were devoid of protein(s) immunoreactive to anti-CYP2A6 antibody. Thus, this individual was suggested to be due to the gene deletion in CYP2A6. By analyzing the remaining 38 Japanese and 43 Caucasians, we found that there were no cases of CYP2A6 * 3-type polymorphism in the samples examined in this study, and no cases of CYP2A6 * 2-type polymorphism in the Japanese samples. Of Caucasians studied two individuals were classified into heterozygous CYP2A6 * 1/ * 2-type. Liver microsomal coumarin 7-hydroxylation activities in these two Caucasians were found to be lower than those of the other 41 Caucasians. Kinetic analysis showed that two CYP2A6 * 1/ * 2 individuals had a very low ratio of V max to K m for nicotine C-oxidation as well as coumarin 7-hydroxylation in liver microsomes, compared with those of homozygous CYP2A6 * 1-type. These results suggest that among 39 Japanese and 43 Caucasians examined one Japanese is classified to be CYP2A6 gene deletion and two Caucasians are heterozygous CYP2A6 * 1/ * 2-genotype. Thus the race-related differences in the occurrence of CYP2A6 genetic polymorphisms were supported.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002040050005
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  • 2
    Electronic Resource
    Electronic Resource
    Roles of CYP2A6 and CYP2B6 in nicotine C-oxidation by human liver microsomes (1999)
    Springer
    Archives of toxicology 73 (1999), S. 65-70 
    Details
    ISSN: 1432-0738
    Keywords: Key words Nicotine ; CYP2A6 ; CYP2B6 ; Human P450 ; Liver microsomes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Nicotine C-oxidation by recombinant human cytochrome P450 (P450 or CYP) enzymes and by human liver microsomes was investigated using a convenient high-performance liquid chromatographic method. Experiments with recombinant human P450 enzymes in baculovirus systems, which co-express human nicotinamide adenine dinucleotide phosphate (reduced form) (NADPH)-P450 reductase, revealed that CYP2A6 had the highest nicotine C-oxidation activities followed by CYP2B6 and CYP2D6; the K m values by these three P450 enzymes were determined to be 11.0, 105, and 132 μM, respectively, and the V max values to be 11.0, 8.2, and 8.6 nmol/min per nmol P450, respectively. CYP2E1, 2C19, 1A2, 2C8, 3A4, 2C9, and 1A1 catalysed nicotine C-oxidation only at high (500 μM) substrate concentration. CYP1B1, 2C18, 3A5, and 4A11 had no measurable activities even at 500 μM nicotine. In liver microsomes of 16 human samples, nicotine C-oxidation activities were correlated with CYP2A6 contents at 10 μM substrate concentration, whereas such correlation coefficients were decreased when the substrate concentration was increased to 500 μM. Contribution of CYP2B6 (as well as CYP2A6) was demonstrated by experiments with the effects of orphenadrine (and also coumarin and anti-CYP2A6) on the nicotine C-oxidation activities by human liver microsomes at 500 μM nicotine. CYP2D6 was found to have minor roles since quinidine did not inhibit microsomal nicotine C-oxidation at both 10 and 500 μM substrate concentrations. These results support the view that CYP2A6 has major roles for nicotine C-oxidation at lower substrate concentration and both CYP2A6 and 2B6 play roles at higher substrate concentrations in human liver microsomes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002040050588
    Library Location Call Number Volume/Issue/Year Availability
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    Paper (German National Licenses)
    Fulltext
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